*Target (protein/gene name): Inositol-phosphate phosphatase *NCBI Gene # or RefSeq#: 5777523 *Protein ID (NP or XP #) or Wolbachia#: YP_001576165.1 *Organism: Staphylococcus aureus Etiologic Risk Group (see link below): Risk Group 2 (RG2) *Background/Disease Information: S. aureus is a facultatively anaerobic, Gram-positive coccus, which appears as grape-like clusters when viewed through a microscope, and has large, round, golden-yellow colonies, often with hemolysis, when grown on blood agar plates.S. aureus can cause a range of illnesses, from minor skin infections, such as pimples, impetigo, boils (furuncles), cellulitis folliculitis, carbuncles, scalded skin syndrome, and abscesses, to life-threatening diseases suchas pneumonia, meningitis, osteomyelitis, endocarditis, toxic shock syndrome (TSS),bacteremia, and sepsis. Its incidence ranges from skin, soft tissue, respiratory, bone, joint, endovascular to wound infections. It is still one of the five most common causes of nosocomial infections and is often the cause of postsurgical wound infections. Each year, some 500,000 patients in American hospitals contract a staphylococcal infection.
Enzyme Assay information (spectrophotometric, coupled assay ?, reagents): http://www.brenda-enzymes.org/literature/lit.php4?e=3.1.3.25&r=713666 -- link to Sigma (or other company) page for assay or assay reagents (substrates) -- link to paper that contains assay information -- List cost and quantity of substrate reagents and supplier
Structure Available (PDB or Homology model) -- PDB # or closest PDB entry if using homology model: 3T0J -- Query Coverage (if not direct match): N/A -- Max Ident (if not direct match): N/A
*NCBI Gene # or RefSeq#: 5777523
*Protein ID (NP or XP #) or Wolbachia#: YP_001576165.1
*Organism: Staphylococcus aureus
Etiologic Risk Group (see link below): Risk Group 2 (RG2)
*Background/Disease Information: S. aureus is a facultatively anaerobic, Gram-positive coccus, which appears as grape-like clusters when viewed through a microscope, and has large, round, golden-yellow colonies, often with hemolysis, when grown on blood agar plates.S. aureus can cause a range of illnesses, from minor skin infections, such as pimples, impetigo, boils (furuncles), cellulitis folliculitis, carbuncles, scalded skin syndrome, and abscesses, to life-threatening diseases suchas pneumonia, meningitis, osteomyelitis, endocarditis, toxic shock syndrome (TSS),bacteremia, and sepsis. Its incidence ranges from skin, soft tissue, respiratory, bone, joint, endovascular to wound infections. It is still one of the five most common causes of nosocomial infections and is often the cause of postsurgical wound infections. Each year, some 500,000 patients in American hospitals contract a staphylococcal infection.
Essentiality of this protein: None according to TDR
Complex of enzymes: None
*EC#: 3.1.3.25
Link to BRENDA EC# page: http://www.brenda-enzymes.org/php/result_flat.php4?ecno=3.1.3.25&Suchword=&organism%5B%5D=Staphylococcus+aureus&show_tm=0
-- Show screenshot of BRENDA enzyme mechanism schematic
Enzyme Assay information (spectrophotometric, coupled assay ?, reagents):
http://www.brenda-enzymes.org/literature/lit.php4?e=3.1.3.25&r=713666
-- link to Sigma (or other company) page for assay or assay reagents (substrates)
-- link to paper that contains assay information
-- List cost and quantity of substrate reagents and supplier
Structure Available (PDB or Homology model)
-- PDB # or closest PDB entry if using homology model: 3T0J
-- Query Coverage (if not direct match): N/A
-- Max Ident (if not direct match): N/A
Druggable Target (see Databases for this): Yes
Current Inhibitors: None
Expression Information (has it been expressed in bacterial cells): Yes, expressed in E. Coli
Purification Method: http://www.brenda-enzymes.org/literature/lit.php4?e=3.1.3.25&r=713666
Image of protein (PyMol or etc):
*Amino Acid Sequence:
MTDKTLQQIDKLICSWLKQIDNVIPQLIMEMTTETKRHRFDLVTNVDKQIQQQFQQFLATHFPEHQLLAE
EKSNEMITNEINHLWIMDPIDGTANLVKQQEDYCIILAYFYEGKPMLSYVYDYPHKKLYKAIRGEGAFCN
GIKMEEPPSLKLEDAIISFNAQVMNLDTVQDLFDASFSYRLVGACGLDSMRVAKGQFGAHINTNPKPWDI
AAQFLFAELLNLKMTTLDGKAIDHLKGAPFIISNKACHETVLKILNANGGYQKYR
*length of your protein in Amino Acids: 265 AA
Molecular Weight of your protein in kiloDaltons using the Expasy ProtParam website: 30385.0 Daltons
*CDS Gene Sequence:
ATGACAGATAAAACATTACAGCAAATAGATAAGTTAATATGTTCGTGGTTGAAACAAATAGACAATGTTA
TTCCGCAATTAATTATGGAAATGACTACAGAAACTAAGCGTCATCGATTTGATTTAGTTACAAATGTAGA
TAAACAGATTCAACAACAATTCCAACAATTTTTAGCAACACATTTTCCGGAACACCAACTATTAGCGGAA
GAAAAAAGTAATGAAATGATTACGAATGAAATTAATCATTTATGGATTATGGATCCCATTGATGGAACTG
CCAATTTAGTAAAACAGCAAGAAGATTATTGTATTATATTGGCGTATTTTTATGAAGGTAAACCAATGTT
ATCGTACGTATATGATTATCCGCATAAAAAGCTTTATAAAGCAATACGAGGAGAAGGTGCTTTTTGTAAT
GGAATTAAGATGGAAGAACCACCATCGTTAAAATTGGAAGACGCTATTATATCGTTTAATGCACAAGTGA
TGAATCTAGATACGGTGCAAGATTTATTTGACGCCTCATTTAGTTATCGTTTAGTTGGTGCTTGCGGATT
AGATTCTATGCGAGTGGCAAAAGGTCAATTTGGGGCACATATTAATACAAATCCTAAACCTTGGGACATT
GCTGCACAGTTTTTATTTGCAGAACTATTGAATCTAAAAATGACGACTTTAGATGGTAAGGCAATTGATC
ATTTAAAGGGTGCACCTTTTATTATAAGTAATAAAGCATGTCATGAAACGGTACTTAAAATTTTAAATGC
AAATGGTGGTTATCAAAAATACAGATAG
*GC% Content for gene: 32%
*CDS Gene Sequence (codon optimized) - copy from output of Primer Design Protocol:
ATGACCGACAAAACCCTGCAGCAGATCGATAAACTCATTTGTTCTTGGCTCAAGCAAATC
GACAATGTGATCCCTCAACTGATTATGGAAATGACGACCGAAACGAAGCGTCACCGTTTC
GACCTCGTTACCAATGTTGACAAACAGATCCAACAGCAGTTCCAGCAATTCCTCGCGACC
CACTTTCCTGAGCACCAGCTGCTGGCGGAAGAAAAATCTAATGAAATGATTACCAACGAA
ATCAACCACCTCTGGATCATGGACCCGATCGACGGTACGGCGAACCTGGTTAAACAACAG
GAAGACTACTGCATCATCCTGGCGTACTTCTACGAAGGTAAACCTATGCTGTCTTACGTT
TATGACTATCCTCACAAAAAACTGTATAAAGCTATCCGCGGTGAGGGTGCATTTTGCAAC
GGTATCAAAATGGAAGAACCGCCTTCTCTCAAACTGGAAGACGCCATCATCTCTTTTAAC
GCGCAGGTTATGAACCTCGACACTGTTCAGGACCTGTTCGATGCGTCCTTTTCTTATCGC
CTGGTTGGTGCGTGCGGTCTGGACTCTATGCGTGTTGCTAAAGGTCAGTTCGGTGCCCAC
ATCAACACCAATCCTAAGCCTTGGGACATCGCAGCGCAATTTCTCTTCGCGGAACTCCTC
AATCTCAAGATGACCACGCTGGACGGTAAAGCCATCGATCACCTGAAAGGCGCTCCGTTC
ATTATTTCTAACAAAGCGTGCCACGAAACCGTTCTGAAAATTCTCAATGCGAACGGTGGT
TACCAGAAATACCGCCTGGGTGACTCTCCGGGTTACGTTATGTCTAACATCGAATTCCGT
CAGCTCACCCGTGGCCATTCTCCATCTGACGAACGTGAAGCTCGCCGTGTCGAAGAAGCC
GGTGGTCAGCTGTTCGTTATTGGTGGTGAACTCCGTGTAAATGGTGTGCTGAACCTGACG
CGTGCCCTGGGCGACGTTCCGGGTCGTCCGATGATCAGCAACGAACCGGAAACCTGTCAA
GTTCCGATTGAGAGCAGCGACTATCTGGTACTCCTGGCATGCGACGGCATCTCTGATGTG
TTCAACGAGCGTGATCTGTACCAACTGGTTGAAGCCTTCGCAAACGACTACCCGGTTGAG
GATTATGCAGAACTGTCTCGTTTCATCTGCACCAAGGCGATCGAGGCTGGTTCTGCGGAC
AACGTCTCTGTTGTGATCGGTTTCCTGCGTCCGCCGCAAGACGTTTGGAAACTGATGAAG
CACGAGTCTGATGACGAGGACTCTGACGTTACCGACGAAGAATAA
*GC% Content for gene (codon optimized): 50%
Primer design results for pNIC-Bsa4 cloning (list seqeunces of all of your ~40 nt long primers):
1 ATGACCGACAAAACCCTGCAGCAGATCGATAAACTCATTTGTTCTTGGCTCAAGC 55
2 TCATTTCCATAATCAGTTGAGGGATCACATTGTCGATTTGCTTGAGCCAAGAACAAATGA 60
3 CCCTCAACTGATTATGGAAATGACGACCGAAACGAAGCGTCACCGTTTCGACCTCGTTAC 60
4 GGAATTGCTGGAACTGCTGTTGGATCTGTTTGTCAACATTGGTAACGAGGTCGAAACGGT 60
5 ACAGCAGTTCCAGCAATTCCTCGCGACCCACTTTCCTGAGCACCAGCTGCTGGCGGAAGA 60
6 TCCAGAGGTGGTTGATTTCGTTGGTAATCATTTCATTAGATTTTTCTTCCGCCAGCAGCT 60
7 CGAAATCAACCACCTCTGGATCATGGACCCGATCGACGGTACGGCGAACCTGGTTAAACA 60
8 CTTCGTAGAAGTACGCCAGGATGATGCAGTAGTCTTCCTGTTGTTTAACCAGGTTCGCCG 60
9 CCTGGCGTACTTCTACGAAGGTAAACCTATGCTGTCTTACGTTTATGACTATCCTCACAA 60
10 ATGCACCCTCACCGCGGATAGCTTTATACAGTTTTTTGTGAGGATAGTCATAAACGTAAG 60
11 CGCGGTGAGGGTGCATTTTGCAACGGTATCAAAATGGAAGAACCGCCTTCTCTCAAACTG 60
12 GGTTCATAACCTGCGCGTTAAAAGAGATGATGGCGTCTTCCAGTTTGAGAGAAGGCGGTT 60
13 ACGCGCAGGTTATGAACCTCGACACTGTTCAGGACCTGTTCGATGCGTCCTTTTCTTATC 60
14 AACACGCATAGAGTCCAGACCGCACGCACCAACCAGGCGATAAGAAAAGGACGCATCGAA 60
15 GTCTGGACTCTATGCGTGTTGCTAAAGGTCAGTTCGGTGCCCACATCAACACCAATCCTA 60
16 GTTCCGCGAAGAGAAATTGCGCTGCGATGTCCCAAGGCTTAGGATTGGTGTTGATGTGGG 60
17 GCAATTTCTCTTCGCGGAACTCCTCAATCTCAAGATGACCACGCTGGACGGTAAAGCCAT 60
18 GCTTTGTTAGAAATAATGAACGGAGCGCCTTTCAGGTGATCGATGGCTTTACCGTCCAGC 60
19 CTCCGTTCATTATTTCTAACAAAGCGTGCCACGAAACCGTTCTGAAAATTCTCAATGCGA 60
20 GGAGAGTCACCCAGGCGGTATTTCTGGTAACCACCGTTCGCATTGAGAATTTTCAGAACG 60
21 CGCCTGGGTGACTCTCCGGGTTACGTTATGTCTAACATCGAATTCCGTCAGCTCACCCGT 60
22 TCTTCGACACGGCGAGCTTCACGTTCGTCAGATGGAGAATGGCCACGGGTGAGCTGACGG 60
23 GCTCGCCGTGTCGAAGAAGCCGGTGGTCAGCTGTTCGTTATTGGTGGTGAACTCCGTGTA 60
24 GGAACGTCGCCCAGGGCACGCGTCAGGTTCAGCACACCATTTACACGGAGTTCACCACCA 60
25 CCCTGGGCGACGTTCCGGGTCGTCCGATGATCAGCAACGAACCGGAAACCTGTCAAGTTC 60
26 TCGCATGCCAGGAGTACCAGATAGTCGCTGCTCTCAATCGGAACTTGACAGGTTTCCGGT 60
27 GGTACTCCTGGCATGCGACGGCATCTCTGATGTGTTCAACGAGCGTGATCTGTACCAACT 60
28 ATAATCCTCAACCGGGTAGTCGTTTGCGAAGGCTTCAACCAGTTGGTACAGATCACGCTC 60
29 GACTACCCGGTTGAGGATTATGCAGAACTGTCTCGTTTCATCTGCACCAAGGCGATCGAG 60
30 GCAGGAAACCGATCACAACAGAGACGTTGTCCGCAGAACCAGCCTCGATCGCCTTGGTGC 60
31 TGTTGTGATCGGTTTCCTGCGTCCGCCGCAAGACGTTTGGAAACTGATGAAGCACGAGTC 60
32 TTATTCTTCGTCGGTAACGTCAGAGTCCTCGTCATCAGACTCGTGCTTCATCAGTTTC 58
https://docs.google.com/file/d/0B4O2KqKh2q_-cWEzcmlETE9MN2s/edit
Primer design results for 'tail' primers (this is just 2 sequences):
Literature:
http://www.ncbi.nlm.nih.gov/pubmed/22360485
http://enzyme.expasy.org/EC/3.1.3.25
http://www.brenda-enzymes.org/php/result_flat.php4?ecno=3.1.3.25&Suchword=&organism%5B%5D=Staphylococcus+aureus&show_tm=0
http://www.brenda-enzymes.org/literature/lit.php4?e=3.1.3.25&r=713666